Molecular Genetic Studies on the Physiological Function and Disorders of DNA Methylation

DNA甲基化生理功能和紊乱的分子遗传学研究

• 批准号: 13307067
• 负责人: SASAKI Hiroyuki
• 金额: $ 34.86万
• 依托单位: National Institute of Genetics
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13307067/
• 关键词: DNA methylation; DNA methyltransferases; methyl-CpG binding proteins; ICF syndrome; Rett syndrome; DNAメチル化; レット症侯群

Methylation of CpG dinucleotides plays a central role in the epigenetic genome regulation, and its defects cause congenital disorders and cancers. We have studied the functions and disorders of DNA methyltransferases (DNMTs) and methyl-CpG-binding proteins (MBD proteins) and found the followings. 1.By knocking out mouse DNMT3A, a de novo-type. DNMT, in a germline-specific manner, we found that this enzyme is essential for paternal and maternal imprinting. 2.Enzymatic properties and target specificities of DNMT3A and DNMT3B were determined. 3.New mutations of DNMT3B were identified in Japanese families with ICE syndrome. We also found an ICE case with no mutation in DNMT3B, which suggests the heterogeneity of the disease. 4.We reported that paternal disomy 14 is a new imprinting-related disorder characterized by bell-shaped chest and wavy ribs. 5.We found that MBD1 represses transcription and forms repressive chromatin through two independent pathways: 1) one mediated by a complex formed with a transcriptional mediator MCAF/AM and a histone methyltransferase SETDB1/ESET; 2) the other involving a histone methyltransferase Suv39, a histone deacetylase HDAC1/2 and a heterochromatin protein HP1. 6.The tertiary structure of the DNA binding domain of MBD1 complexed with a. methylated-CpG-containing DNA molecule was determined. 7.The mutations of McCP2 causing Rett syndrome were found to impair its ability to repress transcription and to form repressive chromatin. 8.We reported that MBD1 promotes base excision repair in cooperation with a methylated-purine DNA glycosylase. All together, our findings have greatly increased our knowledge on the mechanisms of DNA methylation and transcriptional repression through it and the pathology of DNA methylation-associated disorders.

CpG二核苷酸甲基化在表观基因组调控中起着核心作用，它的缺陷会导致先天性疾病和癌症。我们对DNA甲基转移酶(DNMT)和甲基CpG结合蛋白(MBD蛋白)的功能和紊乱进行了研究，发现了以下两种蛋白。1.通过敲除小鼠DNMT3A，一种新的类型。DNMT，以种系特异性的方式，我们发现这种酶对父亲和母亲的印记是必不可少的。2.确定了DNMT3A和DNMT3B的酶学性质和靶向性。3.在日本ICE综合征家系中发现了新的Dnmt3b突变。我们还发现了一例没有Dnmt3b突变的ICE病例，这表明该疾病具有异质性。4.我们报道了父系14号二体是一种新的印记相关疾病，其特征是胸廓呈钟形，肋骨呈波浪形。5.我们发现MBD1通过两条独立的途径抑制转录并形成抑制性染色质：1)由转录介体MCAF/AM和组蛋白甲基转移酶SETDB1/ESET形成的复合体介导；2)组蛋白甲基转移酶Suv39、组蛋白去乙酰化酶HDAC1/2和异染色质蛋白HP1介导的抑制染色质形成。6.测定了含甲基化CpG的DNA分子与MBD1DNA结合区的三级结构。7.发现引起Rett综合征的McCP2突变削弱了其抑制转录和形成抑制性染色质的能力。8.我们报道了MBD1通过与甲基化的嘌呤DNA糖基酶协同促进碱基切除修复。总之，我们的发现极大地增加了我们对DNA甲基化和转录抑制的机制以及DNA甲基化相关疾病的病理学的了解。

项目成果

Hata, K.: "Genomic imprinting : mechanisms, significance and evolution."J.Mamm.Ova.Res.. 20. 64-68 (2003)

Hata, K.：“基因组印记：机制、意义和进化。”J.Mamm.Ova.Res.. 20. 64-68 (2003)

Kudo, S.: "Functional characterization of McCP2 mutations found in male patients with X-linked mental retardation."J.Med.Genet.. 39. 132-136 (2002)

Kudo, S.：“X 连锁智力障碍男性患者中发现的 McCP2 突变的功能特征。”J.Med.Genet.. 39. 132-136 (2002)

Fujita, N.: "Methyl-CpG binding domain 1(MBD1) interacts with Suv39h1-HP1 heterochromatic complex for DNA methylation-based transcriptional repression."J.Biol.Chem.. 278. 24132-24138 (2003)

Fujita, N.：“甲基-CpG 结合域 1 (MBD1) 与 Suv39h1-HP1 异色复合物相互作用，实现基于 DNA 甲基化的转录抑制。”J.Biol.Chem.. 278. 24132-24138 (2003)

Kurosawa, k.: "Paternal UPD14 is responsible for a distinctive malformation complex."Am.J.Med.Genet.. 110. 268-272 (2002)

Kurosawa, k.：“父系 UPD14 导致独特的畸形复合体。”Am.J.Med.Genet.. 110. 268-272 (2002)

Kagotani, k.: "Visualization of transcription-dependent association of imprinted genes with the nuclear matrix."ExCell Res.. 274. 189-196 (2002)

Kagotani, k.：“印记基因与核基质的转录依赖性关联的可视化。”ExCell Res.. 274. 189-196 (2002)